Orpinomyces xylanase proteins and coding sequences

Abstract

Xylanases having high specific activities from Orpinomyces sp. strain PC-2 are provided as well as methods for their purification. DNA sequences encoding these proteins are also provided.

We claim: 
     
       1. An isolated recombinant DNA molecule comprising a nucleotide sequence encoding a xylanase protein from Orpinomyces, wherein said nucleotide sequence is at least 90% homologous to a nucleotide sequence as given in SEQ ID NO:1 nucleotides 169-1185. 
     
     
       2. The isolated recombinant DNA molecule of claim 1 wherein said nucleotide sequence encodes a XynA-2 protein that has an amino acid sequence as given in SEQ ID NO:2 from amino acid 25 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       3. The isolated recombinant DNA molecule of claim 2 wherein said XynA-2 protein is encoded by the nucleotide sequence as given in SEQ ID NO:1 from about nucleotide 169 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       4. The isolated recombinant DNA molecule of claim 2 wherein said XynA-2 protein has an amino acid sequence as given in SEQ ID NO:2 from amino acid 20 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       5. The isolated recombinant DNA molecule of claim 4 wherein said XynA-2 protein is encoded by the nucleotide sequence as given in SEQ ID NO:1 from nucleotide 154 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       6. The isolated recombinant DNA molecule of claim 2 wherein said XynA-2 protein has an amino acid sequence as given in SEQ ID NO:2 from amino acid 15 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       7. The isolated recombinant DNA molecule of claim 6 wherein said XynA-2 protein is encoded by the nucleotide sequence as given in SEQ ID NO:1 from nucleotide 139 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       8. The isolated recombinant DNA molecule of claim 2 wherein said XynA-2 protein has an amino acid sequence as given in SEQ ID NO:2 from amino acid 1 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       9. The isolated recombinant DNA molecule of claim 8 wherein said XynA-2 protein is encoded by the nucleotide sequence as given in SEQ ID NO:1 from nucleotide 97 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       10. The isolated recombinant DNA molecule of claim 2 additionally comprising DNA encoding a signal peptide immediately upstream of and operably linked to the nucleotide sequence encoding said XynA-2 protein. 
     
     
       11. The isolated recombinant DNA molecule of claim 10 wherein said signal peptide is an Aureobasidium pullulans xylanase II (APX-II) signal peptide. 
     
     
       12. An Escherichia coli cell comprising the recombinant DNA molecule of claim 2. 
     
     
       13. A Saccharomyces cerevisiae cell comprising the recombinant DNA molecule of claim 2. 
     
     
       14. An Aspergillus nidulans cell comprising the recombinant DNA molecule of claim 2. 
     
     
       15. A Trichoderma reesei cell comprising the recombinant DNA molecule of claim 2. 
     
     
       16. A recombinant host cell comprising the recombinant DNA molecule of claim 2 wherein said host cell is selected from the group consisting of Pichia, Streptomyces and Bacillus. 
     
     
       17. A method of using a DNA molecule comprising a nucleotide sequence encoding a xylanase protein from Orpinomyces to produce the xylanase protein in a host cell other than Orpinomyces sp. strain PC-2, wherein said nucleotide sequence is at least 90% homologous to a nucleotide sequence as given by SEQ ID NO:1, nucleotides 169-1185, said method comprising the steps of: (a) infecting or transforming said host cell capable of expressing a xylanase coding region with a vector comprising a promoter active in said host cell wherein said promoter is operably linked to the coding region for said xylanase; and   (b) culturing the infected or transformed cell under conditions suitable for expression of said xylanase.   
     
     
       18. The method of claim 17 wherein said host cell is an Escherichia coli cell. 
     
     
       19. The method of claim 17 wherein said host cell is a Saccharomyces cerevisiae cell. 
     
     
       20. The method of claim 17 wherein said host cell is an Aspergillus nidulans cell. 
     
     
       21. The method of claim 17 wherein said host cell is a Trichoderma reesei cell. 
     
     
       22. The method of claim 17 wherein said host cell is selected from the group consisting of Pichia, Streptomyces and Bacillus. 
     
     
       23. The method of claim 17 wherein said vector further comprises a nucleotide sequence encoding a signal peptide operably linked between said promoter and said coding region. 
     
     
       24. The method of claim 23 wherein said peptide is an Aureobasidium pullulans xylanase II (APX-II) signal peptide. 
     
     
       25. The method of claim 17 wherein said DNA molecule encodes a xylanase XynA-2 protein having an amino acid sequence as set forth in SEQ ID NO:2 from amino acid 25 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       26. The method of claim 17 wherein said coding region for said xylanase XynA-2 protein has the nucleotide sequence as set forth in SEQ ID NO:1 from nucleotide 169 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       27. The method of claim 17 wherein said DNA molecule encodes a xylanase XynA-2 protein having an amino acid sequence as set forth in SEQ ID NO:2 from amino acid 20 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       28. The method of claim 17 wherein said coding region for said xylanase XynA-2 protein has the nucleotide sequence as set forth in SEQ ID NO:1 from nucleotide 154 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       29. The method of claim 17 wherein said DNA molecule encodes a xylanase XynA-2 protein having an amino acid sequence as set forth in SEQ ID NO:2 from amino acid 15 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       30. The method of claim 17 wherein said coding region for said xylanase XynA-2 protein has the nucleotide sequence as set forth in SEQ ID NO:1 from nucleotide 139 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       31. The method of claim 17 wherein said DNA molecule encodes a xylanase XynA-2 protein having an amino acid sequence as set forth in SEQ ID NO:2 from amino acid 1 through amino acid 362 or a functionally equivalent sequence with at least 90% homology thereto. 
     
     
       32. The method of claim 17 wherein said coding region for said xylanase XynA-2 protein has the nucleotide sequence as set forth in SEQ ID NO:1 from nucleotide 97 through nucleotide 1185 or a functionally equivalent sequence with at least 90% homology thereto.